1. Field of the Invention
This invention relates broadly to liquid chromatographic apparatus and methods. More particularly, it concerns improved optical systems and methods for measuring various properties of the effluent stream of a liquid chromatographic column.
2. Description of the Prior Art
Liquid chromatography procedures are used to separate an unknown sample into its various chemical components in an elongated column packed with selective adsorbent material and to then make various types of measurements on the effluent stream from the column as to changes in certain properties of the effluent steam relative to time. Hence, in such procedures it is necessary to accurately detect and measure, on a continuous basis, the particular property under observation, e.g., refractive index, light absorbance, etc.
Ultraviolet and visible light absorption detectors are the most widely used detectors in high performance liquid chromatography and have had a significant effect on the development of such chromatography. However, improved detectors also have been developed that measure refractive index change. For example, U.S. Pat. No. 3,950,104 discusses various types of R.I. detectors and discloses and claims improved devices of one of the general types. It also describes flow and refractive index effects in small flow cells and its disclosure is incorporated herein by reference.
Most UV and visible light absorption detectors for liquid chromatography employ flat parallel transparent windows and are limited in the degree of convergence of the light illuminating the flow cell to avoid serious loss in linear dynamic range due to unequal light pathlengths through the cell. Other light absorption detectors employ a cylindrical cell with transverse illumination by more or less parallel light and also provide limited linear dynamic response range due to unequal pathlengths of light through the flow cell. The nonlinearity of response of these detectors is a consequence of the logarithmic dependence of the transmitted light on sample concentration in accordance with Beer's Law, i.e., log (I.sub.o /I)=a.times.b.times.c, where I.sub.o is the light incident on the flow cell, I is the light transmitted through the flow cell, a is the molar absorptivity, b is the pathlength and c is the molar sample concentration in the flow cell.
Improved detectors for chromatography have been developed for achieving a high linear dynamic response range in cylindrical cells with transverse illumination (see U.S. Pat. Nos. 3,975,104 and 4,006,990)
U.S. Pat. No. 3,975,104 describes the use of a slit aperture along the axis of the transversely illuminated flow cell to limit the light rays used in the sample concentration measurement to those which pass near the axis of the flow cell, that are therefore of nearly equal pathlength in the cell. However, the slit aperture within the cell complicates construction of the cell and perturbs the cylindrical symmetry of the flow through the cell. Slit and circular apertures have been used in a multitude of other ways in a variety of other optical systems (see for example U.S. Pat. Nos. 2,540,827; 3,075,426 and 3,999,856).
U.S. Pat. No. 4,006,990 describes the use of parallel light in conjunction with a positive lens ahead of the flow cell to focus the light near the center of the flow cell. The light rays converging near the center of the cell have nearly equal pathlengths through the flow cell. This means of obtaining nearly equal pathlengths of light in the cell requires a source of parallel light such as that obtained in expensive spectrophotometer optics or lasers.
Although there has been much research and development work on optical systems for use in liquid chromatography as indicated by the references cited above, there is a need for improved means for providing nearly equal pathlengths in transversely illuminated cylindrical flow cells in which construction of the flow cell would be simpler than in the known systems and which would provide axial symmetry of fluid flow through the cell. This invention concerns improved equipment that provides these needs while retaining all the essential advantages of the prior art.